Aurora-A kinase-inactive mutants disrupt the interaction with Ajuba and cause defects in mitotic spindle formation and G2/M phase arrest in HeLa cells

Coordination of cell growth and proliferation in response to nutrient supply is mediated by mammalian target of rapamycin (mTOR) signaling. In this study, we report that Mio, a highly conserved member of the SEACAT/GATOR2 complex necessary for the activation of mTORC1 kinase, plays a critical role in mitotic spindle formation and subsequent chromosome segregation by regulating the proper concentration of active key mitotic kinases Plk1 and Aurora A at centrosomes and spindle poles. Mio-depleted cells showed reduced activation of Plk1 and Aurora A kinase at spindle poles and an impaired localization of MCAK and HURP, two key regulators of mitotic spindle formation and known substrates of Aurora A kinase, resulting in spindle assembly and cytokinesis defects. Our results indicate that a major function of Mio in mitosis is to regulate the activation/deactivation of Plk1 and Aurora A, possibly by linking them to mTOR signaling in a pathway to promote faithful mitotic progression.

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Nucleophosmin is required for chromosome congression, proper mitotic spindle formation, and kinetochore-microtubule attachment in HeLa cells

FEBS Letters ◽

10.1016/j.febslet.2008.10.023 ◽

2008 ◽

Vol 582 (27) ◽

pp. 3839-3844 ◽

Cited By ~ 38

Author(s):

Mohammed Abdullahel Amin ◽

Sachihiro Matsunaga ◽

Susumu Uchiyama ◽

Kiichi Fukui

Keyword(s):

Hela Cells ◽

Mitotic Spindle ◽

Spindle Formation ◽

Microtubule Attachment ◽

Chromosome Congression

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Glucocappasalin Induces G2/M-Phase Arrest, Apoptosis, and Autophagy Pathways by Targeting CDK1 and PLK1 in Cervical Carcinoma Cells

Frontiers in Pharmacology ◽

10.3389/fphar.2021.671138 ◽

2021 ◽

Vol 12 ◽

Author(s):

Guangya Xu ◽

Xueling Yan ◽

Zhongjia Hu ◽

Lulu Zheng ◽

Ke Ding ◽

...

Keyword(s):

Cell Cycle ◽

Cervical Cancer ◽

Cervical Carcinoma ◽

Hela Cells ◽

Carcinoma Cells ◽

Dependent Manner ◽

Cycle Arrest ◽

Phase Arrest ◽

M Phase

Glucocappasalin (GCP), a natural product derived from the seeds of Descurainia sophia (L.) Webb. ex Prantl, exhibits potential antitumor activity in HeLa cervical carcinoma cells. In this study, we investigated the anti-cervical cancer property of GCP through the induction of cell cycle arrest, apoptosis, and autophagy in vitro and in vivo, and elucidated the underlying molecular mechanisms. We demonstrated that treatment with GCP inhibited the growth of HeLa, Siha, and Ca Ski cell lines in a dose-dependent manner, with HeLa cells displaying particular sensitivity to the GCP treatment. Subsequently, the expression of cyclin-dependent kinase 1 (CDK1) and polo like kinase 1 (PLK1) were evaluated in HeLa cells using the CDK1 kinase assay kit, the fluorescence polarization assay, real-time quantitative PCR, and western blotting. Our results demonstrate that GCP could be employed to attenuate the expression of CDK1 and PLK1 in a dose- and time-dependent manner. The complementary results obtained by flow cytometry and western blotting allowed us to postulate that GCP may exhibit its antitumor effects by inducing G2/M cell cycle arrest. Moreover, HeLa cells treated with GCP exhibited a loss in mitochondrial membrane potential, together with the activation of caspases 3 and 9, and poly ADP-ribose polymerase (PARP). Additionally, we found that GCP could increase the formation of acidic vesicular organelles (AVOs), as well as the levels of Beclin1, LC3-II, p62, and Atg5 proteins in HeLa cells. Further studies indicated that GCP triggered autophagy via the suppression of the PI3K/AKT/mTOR signaling pathways. The autophagy inhibitor 3-methyladenine (3-MA) was used to determine whether autophagy affects the apoptosis induced by GCP. Interestingly, the inhibition of autophagy attenuated apoptosis. In vivo anti-tumor experiments indicated that GCP (60 mg/kg, i.p.) markedly reduced the growth of HeLa xenografts in nude mice without apparent toxicity. Taken together, we demonstrate that GCP induces cell cycle G2/M-phase arrest, apoptosis, and autophagy by acting on the PI3K/AKT/mTOR signaling pathways in cervical carcinoma cells. Thus, GCP may represent a promising agent in the eradication of cervical cancer.

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A Novel Aurora-a Kinase Inhibitor MLN8237 Induces Cytotoxicity and Cell Cycle Arrest in Experimental Multiple Myeloma Models.

Blood ◽

10.1182/blood.v112.11.1719.1719 ◽

2008 ◽

Vol 112 (11) ◽

pp. 1719-1719 ◽

Cited By ~ 1

Author(s):

Gullu Gorgun ◽

Elisabetta Calabrese ◽

Mala Mani ◽

Teru Hideshima ◽

Hiroshi Ikeda ◽

...

Keyword(s):

Cell Cycle ◽

Multiple Myeloma ◽

Cell Lines ◽

Mitotic Spindle ◽

Kinase Inhibitor ◽

Thymidine Incorporation ◽

Significant Inhibition ◽

Aurora A Kinase ◽

Aurora A ◽

Resistant Line

Abstract Multiple myeloma (MM) is an incurable bone marrow derived plasma cell malignancy. Despite significant improvements in treating patients suffering from this disease, MM remains uniformly fatal owing to intrinsic or acquired drug resistance. Thus, additional modalities for treating MM are required. In this study, we examined the anti-tumor activity of MLN8237, a small molecule Aurora-A kinase inhibitor, in experimental models of MM. Aurora-A is a mitotic kinase that localizes to centrosomes and the proximal mitotic spindle and functions in mitotic spindle formation and in regulating chromatid congression and segregation. Aurora-A gene amplification and protein overexpression is a common event in many cancers, and has been experimentally linked to genetic instability and tumorigenesis. In MM, increased Aurora-A gene expression has previously been correlated with centrosome amplification and a worsened disease prognosis. Thus, inhibition of Aurora A in MM may prove to be therapeutically beneficial. Here we show that Aurora-A protein is highly expressed in eight distinct MM cell lines. The affect of Aurora-A inhibition in these cell lines was examined in cytotoxicity (MTT viability) and proliferation (3[H]thymidine incorporation) assays by treating with MLN8237 (0.25 mM −32 mM) for 24, 48 and 72h. Although there was no significant inhibition of cell viability and proliferation at 24h, a marked effect occurred 48 and 72h after compound addition at concentrations as low as 0.25 mM. Interestingly, the melphalan resistant line (LR5) and Doxorubucin resistant line (Dox40) were among the least sensitive to MLN8237 induced cell cytotoxicity. The affect of MLN8237 on peripheral blood mononuclear cells (PBMCs) from healthy donors was also examined at the same concentrations and exposure time used for the MM cell lines. In healthy PBMCs, MLN8237 did not induce cytotoxicity as measured by the MTT assay, but there was a significant inhibition of proliferation at 48 and 72h as measured by the 3[H]thymidine incorporation assay at concentrations above 4uM. To delineate the mechanisms of cytotoxicity and growth inhibitory activity of MLN8237, apoptotic markers and cell cycle profiles were examined in the MM cell lines. Fluorescence conjugated-Annexin V and propidium iodide (PI) co-staining of MM cell lines after culturing in the presence or absence of MLN8237 at 1 mM (IC50) for 24, 48 and 72h demonstrated that MLN8237 induces apoptosis in these lines. This finding was corroborated by demonstrating increased capase-9 expression by Western blot analysis. Cell cycle analysis by flow cytometry demonstrated that MLN8237 results in an accumulation of tetraploid cells, presumably by abrogating G2/M progression. These results suggest that MLN8237 represents a possible novel agent for treating MM patients. Additional studies are ongoing to assess the anti-tumor effects of MLN8237 alone and in combination with other therapeutic agents in xenograft models of MM.

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Polypeptide Fraction fromArca subcrenataInduces Apoptosis and G2/M Phase Arrest in HeLa Cells via ROS-Mediated MAPKs Pathways

Evidence-based Complementary and Alternative Medicine ◽

10.1155/2015/930249 ◽

2015 ◽

Vol 2015 ◽

pp. 1-12 ◽

Cited By ~ 4

Author(s):

Xianjing Hu ◽

Zhang Zhang ◽

Ting Liu ◽

Liyan Song ◽

Jianhua Zhu ◽

...

Keyword(s):

Cervical Cancer ◽

Hela Cells ◽

Underlying Mechanism ◽

Cyclin B1 ◽

Mitochondrial Pathway ◽

Dependent Manner ◽

Phase Arrest ◽

M Phase ◽

Induced Apoptosis ◽

High Level

Arca subcrenatais documented in the literature of marine Traditional Chinese Medicine. Polypeptide fraction fromA. subcrenata, coded as P2, was demonstrated to possess significant antitumor activity in our previous study. However, the underlying mechanism remains undefined. The present study was carried out to investigate the underlying antitumor mechanism of P2 in human cervical cancer HeLa cells by MTT, FCM, LSCM, and western blot assays. The results revealed that P2 significantly induced apoptosis of HeLa cells in a concentration- and time-dependent manner. High level of ROS was provoked by P2, which was in turn responsible for induction of apoptosis through activation of intrinsic mitochondrial pathway and JNK1/2, p38 MAPK pathways, as well as inhibition of ERK1/2 pathway, as evidenced by the abrogation of P2’s effect on HeLa cells preincubated with the ROS scavenger NAC. P2 also was observed to display significant effect on G2/M phase arrest by downregulating the expression of cyclin B1/cdc2 complex and upregulating the expression of p21. These findings demonstrate that P2 induces apoptosis and G2/M phase arrest in HeLa cells through ROS-mediated MAPKs pathways, suggesting that P2 would be worth investigating as a promising agent within the scope of marine drugs for treatment of cervical cancer.

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Kaempferol-7-O-β-d-glucoside (KG) isolated from Smilax china L. rhizome induces G2/M phase arrest and apoptosis on HeLa cells in a p53-independent manner

Cancer Letters ◽

10.1016/j.canlet.2008.01.044 ◽

2008 ◽

Vol 264 (2) ◽

pp. 229-240 ◽

Cited By ~ 61

Author(s):

Wen Xu ◽

Jianwen Liu ◽

Changlong Li ◽

He-Zhen Wu ◽

Yan-Wen Liu

Keyword(s):

Hela Cells ◽

Independent Manner ◽

Phase Arrest ◽

M Phase

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Oxidative stress induces mitotic arrest by inhibiting Aurora A-involved mitotic spindle formation

Free Radical Biology and Medicine ◽

10.1016/j.freeradbiomed.2016.12.031 ◽

2017 ◽

Vol 103 ◽

pp. 177-187 ◽

Cited By ~ 18

Author(s):

Guang-Fei Wang ◽

Qincai Dong ◽

Yuanyuan Bai ◽

Jing Yuan ◽

Quanbin Xu ◽

...

Keyword(s):

Oxidative Stress ◽

Mitotic Spindle ◽

Mitotic Arrest ◽

Aurora A ◽

Spindle Formation

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Correction: Microwave-assisted synthesis of polypyridyl ruthenium(ii) complexes as potential tumor-targeting inhibitors against the migration and invasion of Hela cells through G2/M phase arrest

RSC Advances ◽

10.1039/c7ra90067h ◽

2017 ◽

Vol 7 (48) ◽

pp. 29925-29925

Author(s):

Jieqiong Cao ◽

Qiong Wu ◽

Wenjie Zheng ◽

Li Li ◽

Wenjie Mei

Keyword(s):

Hela Cells ◽

Tumor Targeting ◽

Migration And Invasion ◽

Microwave Assisted Synthesis ◽

Microwave Assisted ◽

Phase Arrest ◽

M Phase

Correction for ‘Microwave-assisted synthesis of polypyridyl ruthenium(ii) complexes as potential tumor-targeting inhibitors against the migration and invasion of Hela cells through G2/M phase arrest’ by Jieqiong Cao et al., RSC Adv., 2017, 7, 26625–26632.

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Centriole-independent mitotic spindle assembly relies on the PCNT–CDK5RAP2 pericentriolar matrix

The Journal of Cell Biology ◽

10.1083/jcb.202006010 ◽

2020 ◽

Vol 219 (12) ◽

Author(s):

Sadanori Watanabe ◽

Franz Meitinger ◽

Andrew K. Shiau ◽

Karen Oegema ◽

Arshad Desai

Keyword(s):

Cell Lines ◽

Cancer Cell ◽

Hela Cells ◽

Mitotic Spindle ◽

Spindle Assembly ◽

Cell Type ◽

Matrix Assembly ◽

Spindle Formation ◽

Pericentriolar Material ◽

Mitotic Spindle Assembly

Centrosomes, composed of centrioles that recruit a pericentriolar material (PCM) matrix assembled from PCNT and CDK5RAP2, catalyze mitotic spindle assembly. Here, we inhibit centriole formation and/or remove PCNT–CDK5RAP2 in RPE1 cells to address their relative contributions to spindle formation. While CDK5RAP2 and PCNT are normally dispensable for spindle formation, they become essential when centrioles are absent. Acentriolar spindle assembly is accompanied by the formation of foci containing PCNT and CDK5RAP2 via a microtubule and Polo-like kinase 1–dependent process. Foci formation and spindle assembly require PCNT-CDK5RAP2–dependent matrix assembly and the ability of CDK5RAP2 to recruit γ-tubulin complexes. Thus, the PCM matrix can self-organize independently of centrioles to generate microtubules for spindle assembly; conversely, an alternative centriole-anchored mechanism supports spindle assembly when the PCM matrix is absent. Extension to three cancer cell lines revealed similar results in HeLa cells, whereas DLD1 and U2OS cells could assemble spindles in the absence of centrioles and PCNT-CDK5RAP2, suggesting cell type variation in spindle assembly mechanisms.

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Microwave-assisted synthesis of polypyridyl ruthenium(ii) complexes as potential tumor-targeting inhibitors against the migration and invasion of Hela cells through G2/M phase arrest

RSC Advances ◽

10.1039/c7ra00658f ◽

2017 ◽

Vol 7 (43) ◽

pp. 26625-26632 ◽

Cited By ~ 9

Author(s):

Jieqiong Cao ◽

Qiong Wu ◽

Wenjie Zheng ◽

Li Li ◽

Wenjie Mei

Keyword(s):

Cancer Cells ◽

Hela Cells ◽

Tumor Targeting ◽

Migration And Invasion ◽

Microwave Assisted Synthesis ◽

Microwave Assisted ◽

Potential Inhibitor ◽

Phase Arrest ◽

M Phase

The polypyridyl ruthenium(ii) complexes 4 was identified as a potential inhibitor against the migration and invasion of Hela cells, which could selectively accumulate in tumors tissue and induce G2/M phase arrest in cancer cells.

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